ABSTRACT: It is well established that the incidence and severity of sepsis are increased in older adults, however, targeting of aging-associated alterations in host immune responses has not been successful in clinical trials. As gastrointestinal flora are the primary source of pathogenic bacteria causative of sepsis in the geriatric population (contributing to pneumonia, urinary tract infections, and abdominal infection), we hypothesized that sepsis severity reflects pathogenic shifts in the aging gut microbiome. The novel concept described in this proposal not only addresses a key knowledge gap in sepsis research, but also offers a new conceptual theory that will be the basis for robust future investigations into the management of sepsis in the aging population. In preliminary studies, we demonstrated aging-induced accentuation of sepsis severity by using cecal ligation and puncture (CLP), a gold standard model of gastrointestinal flora-induced sepsis. CLP-induced sepsis severity was augmented in aged mice, yielding increased evidence of systemic inflammation, organ injury, and mortality. Strikingly, this aging-associated augmentation of severity could be replicated in young mice by the intraperitoneal injection of aged (but not young) stool. These findings suggest that the aged microbiome (inclusive of microbiota and associated virulence factors) is the key driver of exaggerated sepsis severity associated with aging. We have also demonstrated microbiota shifts towards known pathogenic genera with aging and that live bacteria are necessary to cause this phenotype. In this proposal, we will rigorously determine microbiota shifts throughout the murine lifespan and correlate these shifts with divergence in fecal slurry induced sepsis severity (Aim 1). Secondly, in collaboration with the Gnotobiotic Core at the University of Colorado, we will experimentally manipulate the gut microbiome of germ- free mice to replicate either young or aged gut microbiome. We will then perform CLP on these animals and measure disease severity (Aim 2). Lastly, we will manipulate the aged gut microbiome in vivo by employing antibiotic therapy targeted for pathogenic genera demonstrated in our microbiome data. We will assess our ability to alter sepsis severity through this pragmatic therapeutic strategy (Aim 3). We believe that the proposed experiments will further our mechanistic understanding of the pathophysiology underlying exaggerated sepsis severity in the aging population and identify novel, feasible, and clinically relevant therapeutic targets. Furthermore, the work proposed in this application will allow Dr. Colbert to develop a unique scientific niche bridging critical illness, host-pathogen interaction, and approaches to microbiome biology.